Channeling of Eukaryotic Diacylglycerol into the Biosynthesis of Plastidial Phosphatidylglycerol

Abstract

Plastidial glycolipids contain diacylglycerol (DAG) moieties, which are either synthesized in the plastids (prokaryotic lipids) or originate in the extraplastidial compartment (eukaryotic lipids) necessitating their transfer into plastids. In contrast, the only phospholipid in plastids, phosphatidylglycerol (PG), contains exclusively prokaryotic DAG backbones. PG contributes in several ways to the functions of chloroplasts, but it is not known to what extent its prokaryotic nature is required to fulfill these tasks. As a first step toward answering this question, we produced transgenic tobacco plants that contain eukaryotic PG in thylakoids. This was achieved by targeting a bacterial DAG kinase into chloroplasts in which the heterologous enzyme was also incorporated into the envelope fraction. From lipid analysis we conclude that the DAG kinase phosphorylated eukaryotic DAG forming phosphatidic acid, which was converted into PG. This resulted in PG with 2-3 times more eukaryotic than prokaryotic DAG backbones. In the newly formed PG the unique ⌬3-trans-double bond, normally confined to 3-transhexadecenoic acid, was also found in sn-2-bound cis-unsaturated C18 fatty acids. In addition, a lipidomics technique allowed the characterization of phosphatidic acid, which is assumed to be derived from eukaryotic DAG precursors in the chloroplasts of the transgenic plants. The differences in lipid composition had only minor effects on measured functions of the photosynthetic apparatus, whereas the most obvious phenotype was a significant reduction in growth.

Activation of the Chloroplast Monogalactosyldiacylglycerol Synthase MGD1 by Phosphatidic Acid and Phosphatidylglycerol

“…It is undetectable in the isolated envelope fraction by conventional techniques. The PA level in the envelope is possibly close to PA levels in thylakoids, which was reported to be 0.08 mol% mainly prokaryotic PA (27). MGD1 was activated by several molecular species of PA from either a prokaryotic or eukaryotic nature.…”

Lipid Biosynthesis and Protein Concentration Respond Uniquely to Phosphate Supply during Leaf Development in Highly Phosphorus-Efficient Hakea prostrata

“…This differs from the situation in Arabidopsis, where levels of all phospholipids show a positive correlation and all glycoglycerolipids show a negative correlation with Pi supply (Dörmann and Benning, 2002). By contrast, 36:x SQDG species, which are synthesized from the ER-derived secondary DAG pool in the chloroplast (Fritz et al, 2007;Li-Beisson et al, 2010;Boudière et al, 2014), clustered in group II (b) and showed the typically documented decrease with increasing Pi supply in H. prostrata. These contrasting responses resulted in an overall sulfolipid profile that responded very little to altered Pi availability.…”

“…In Arabidopsis leaves, 34:x lipid species show mixed acyl chain configurations suggesting biosynthesis from both ER-derived and chloroplast DAG pools, making it hard to determine their precise origin. In Arabidopsis leaves, PG 34:4, SQDG 34:3/34:4, MGDG 34:4 to 34:6, and DGDG 34:1 species almost exclusively carried the C-16 fatty acid in the sn-2 position (DGDG 34:4 species were not detected; Devaiah et al, 2006;Maatta et al, 2012), which would suggest that these lipid species are synthesized from the primary DAG pool in the chloroplast inner envelope membrane via the so-called prokaryotic pathway (Fritz et al, 2007;Li-Beisson et al, 2010;Boudière et al, 2014). In Arabidopsis, both PG and sulfolipids are essential for chloroplast structure and function, but only about 30% of PG species are actually produced in the chloroplast Yu and Benning, 2003).…”

“…PG and SQDG production in the chloroplast are tightly regulated in an antagonist fashion (Essigmann et al, 1998). The fact that H. prostrata appears to produce most of its PG in the chloroplast might account for the fact that only the sulfolipid (and galactolipid) species connected to the primary DAG pool of the chloroplast show the typical compensatory response to lower PG levels in mature leaves (Fritz et al, 2007;Shimojima et al, 2009;Boudière et al, 2014), despite their overall higher P status. The inferred contribution of the prokaryotic pathway for MGDG synthesis rose to as much as 20% in leaves with toxicity symptoms compared with about 2% in leaves of P-limited plants (Supplemental Table S3).…”

Role of phosphatidic acid in plant galactolipid synthesis

“…The plasma membrane is enriched in PA (3e4 mol % of lipids [11]) whereas, in the chloroplast membranes, PA is very low, never detected in the chloroplast envelope, and around 0.1 mol % of lipids in tobacco thylakoids as reported by [12]. The authors detected mainly a form of PA with C18:1 at sn-1 and C16:0 at sn-2 position of glycerol, which is considered as a prokaryotic form of glycerolipid typically synthesized in chloroplasts as we will see below.…”